The nanotoxicity is one of the main issues of Bionanoscience due to possible serve injury to to human health. Especially, silver ion (Ag+) is a representative toxicity metal ion. Silver compounds have been widely employed in industrial production such as electronics, photography and mirrors. In the rapid growth of silver ion exposure, the sensitive detection of silver ions has attracted considerable attention. In this study, we propose the de novo sensing method for silver ions via Kelvin probe force microscope (KPFM). In this method, 30 base pairs of cytosine have been used for silver-specific DNA that is able to compose the cytosine-Ag+-cytosine (C-Ag+-C) conjugates. Because of surface potential difference, KPFM is able to separate the silver ion binding.